• Korean Journal of Microbiology
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• v.35 no.3
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• pp.218-225
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• 1999

We isolated the ANRI fragment from Aspergillus nidulons that could autononlously replicate and enhance transformation efficiency about $10^4$ fold compared lo the integrative vector in Saccha,omgcer cerevisioe. In A. nidulans recombinant plasmid pLJ16-4.5 which carries the 4.5 kb EcoRI fragment of ANRI showed a 170-[old increase of transformation efficiency compared to the integrative vector pLJ16 and could be recovered from iransfonnants as an intact form. Estimated copy number of transforming plasmid pLJ16-4.5 was scored as 2 to 3 copies in transformed A. nidulans. Recoinbinant plasinid pILJ16-4.5 is inilotically unstable; being lost Irom 65% of aswual progeny of transformants on selective medium and 90% on complete medium. Southern analysis of transformant DNA showed that the pILJ16-4.5 is maintained in free form. The sequencing data showed that ANRl fragment was originated from mitochondiral DNA of A. nid~ilans and contained high AT content as much as 74.7%. One ARS consensus sequence (A/T)TTr4T(A/G)TTT(AiT). I I ARS-like sequence (agreement 10 of 11) and ABFl binding core consensus sequence (TCN7ACG). Also six gyrase binding core consensus sequence (YRTGNYNNY: y=C or T, R=A or G, N=A, G, C or T) of $\Phi$X174 and SV40 DNA and one b site (CACTTTACC) combining with gyrase in ColEl are shown. ANRl can be developed as a repl&ng plasinid for lransfoimation system in A. nirlulmis.

• The Korean Journal of Mycology
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• v.40 no.4
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• pp.224-228
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• 2012

In order to screen interactor(s) of the Aspergillus nidulans ${\alpha}$-COP of COPI vesicle, we performed the yeast two hybrid screening by using the gene for A. nidulans ${\alpha}$-COP as a bait and identified ${\varepsilon}$-COP of the COPI vesicle as an interacting protein. The A. nidualns gene for the ${\varepsilon}$-COP was designated $aneA^+$ ($\underline{A.}$ $\underline{n}$idulans $\underline{e}$psi-lone-COP), which encoded 296 amino acid residues with high level of identity with orthologs from other fungi. Domain analyses with yeast two-hybrid system suggested that the interaction between ${\alpha}$-COP and ${\varepsilon}$-COP relied on the C-terminus of both proteins, and that the N-terminal WD domian of ${\alpha}$-COP and the TPR region of ${\varepsilon}$-COP were not essential but required for the enhancement of the interaction. These results indicate that the interaction mode between ${\alpha}$-COP and ${\varepsilon}$-COP of COPI vesicle is evolutionarily well conserved in eukaryotes.

• Journal of Microbiology
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• v.39 no.2
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• pp.95-101
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• 2001

In an effort to develop an efficient expression and secretion system for heterologous proteins in Aspergilius nidulans, the PCR-amplified coding sequence for alkaline pretense (AlpA) of A. oryzae was cloned into a fungal expression vector downstream of A. nidulans aicA (alcohol dehydrogenase) promoter to yield pRAAlp. Transformation of A. nidulans with pRAAlp gave stable transformants harboring various copy numbers (3 to 10) of integrated alpA gene, from among which 6 representatives were selected. On a medium containing 0.8% ammonium sulfate that represses the expression of the host's own pretense, the alcA prumoter-controlled AlpA expression was strongly induced by threonine but repressed by glucose. The level of AlpA secretion was highest (approximately 666 mU/ml) in transformant ALP6 containing the largest copy number integrated alpA. However, the level of AlpA secretion was not necessarily proportional to the copy numbers of the integrated alpA genes. The N-terminal sequence or the secreted mature AlpA was determined to be Gly-Leu-Thr-Thr-Gln-Lys-Ser and its molecular mass to be approximately 34 kDa, indicating that AlpA is properly processed by the removal of 121 N-terminal amino acids.

• Microbiology and Biotechnology Letters
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• v.36 no.3
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• pp.189-194
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• 2008

The phospholipase B (PLB) families are enzymes sharing phospholipase (PL), lysophospholipase (LPL) and lysophospholipase-transacylase (LPTA) activities. In this study, we report the putative gene encoding phospholipase B (plbA) containing lipase motifs was cloned for the first time from the filamentous fungus, Aspergillus nidulans. plbA was isolated from A. nidulans genomic DNA library using a PCR-amplified probe, which is designed on the basis of sequence information derived from the conserved lipase regions of various PLBs. The deduced product of plbA is of 626 amino acids. From the assigned sequence, PlbA showed 72% identity with Penicillium notatum PLB but have low similarity with phospholipase A of other organisms.

• Journal of Environmental Science International
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• v.17 no.11
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• pp.1221-1226
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• 2008

A variety of fungal species are known to degrade cyanide through the action of cyanide hydratase, a specialized nitrilases which hydrolyze cyanide to formamide. This work is a report on two unknown and un-characterized members from Neurospora crassa and Aspergillus nidulans. Recombinant forms of three cyanide hydratases (CHT) originated from N. crassa, Gibberella zeae, and A. nidulans were prepared after their genes were cloned with N-terminal hexahistidine purification tags, expressed in E. coli and purified using immobilized metal affinity chromatography. These enzymes were compared according to their pH activity profiles, and kinetic parameters. Although all three were similar, the N. crassa CHT has the widest pH range of activity above 50% and highest turnover rate ($6.6{\times}10^8min^{-1}$) among them. The CHT of A. nidulans has the highest Km value of the three nitrilases evaluated in here. Expression of CHT in both N. crassa and A. nidulans were induced by the presence of KCN, regardless of any presence of nitrogen sources. These data can be used to determine optimal procedures for the enzyme uses in the remediation of cyanide-containing wastes.

• The Journal of Natural Sciences
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• v.11 no.1
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• pp.45-54
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• 1999

In epistatic grouping of uvs genes in A. nidulans based on the sensitivities to 4-NQO, the same epistatic grouping was obtained as those for UV and MMS-sensitivities. Based on the MMS-sensitivities, uvsA demonstrated synergistic interactions to uvsF and uvsH, the UvsF group genes, but exhibited epistatic interactions to uvsB and uvsC. The same epistatic grouping was also seen for uvsI when UV was irradiated after 4h germination of conidia, showing synergistic interactions to uvsH, uvsC, and uvsB. However, epistatic interactions were observed with uvsF, which were different from those obtained in quiescent conidia by UV. Intergenic and intragenic recombination frequencies were normal in uvsI compared with wild type.

• Journal of Microbiology and Biotechnology
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• v.31 no.5
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• pp.676-685
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• 2021

RNA-binding proteins are involved in RNA metabolism and posttranscriptional regulation of various fundamental biological processes. The PUF family of RNA-binding proteins is highly conserved in eukaryotes, and its members regulate gene expression, mitochondrial biogenesis, and RNA processing. However, their biological functions in Aspergillus species remain mostly unknown in filamentous fungi. Here we have characterized the puf genes in the model organism Aspergillus nidulans. We generated deletion mutant strains for the five putative puf genes present in the A. nidulans genome and investigated their developmental phenotypes. Deletion of pufA or pufE affected fungal growth and asexual development. pufA mutants exhibited decreased production of asexual spores and reduced mRNA expression of genes regulating asexual development. The pufE deletion reduced colony growth, increased formation of asexual spores, and delayed production of sexual fruiting bodies. In addition, the absence of pufE reduced both sterigmatocystin production and the mRNA levels of genes in the sterigmatocystin cluster. Finally, pufE deletion mutants showed reduced trehalose production and lower resistance to thermal stress. Overall, these results demonstrate that PufA and PufE play roles in the development and sterigmatocystin metabolism in A. nidulans.

• Journal of Microbiology
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• v.34 no.1
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• pp.25-29
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• 1996

Since sequencing of randomly selected cDNA clones has been known to be a powerful approach to obtain information on gene expression pattern in specific cells or tissues, we have analyzed a 3'-directed cDNA library of vegetative mycelia of A. nidulans by single-pass sequencing of hundreds of randomly selected clones. Sequencing of 292 cDNA clones yielded 209 gene signatures (GSs) probably representing highly or lesser expressed genes in the vegetative mycelia. Among the 209 GSs, 25 (79 cDNA clones) appeared more than once and 184 only once. One GS appeared at a highest frequency of 6 times, 2 GSs5 times, 4 GSs 4 times, a GSs 3 times and 16 GSs twice. About 6.6% GSs comprizing of 13 GSs showed alternative polyadenylation. Among 23 redundant GSs, three were common in both mycelia and sexual organs, and 22 were probably mycelia-specific. Out of 209 GSs, 36 were identified in GenBank showing of 70% or greater similaritis. Only six GSs were for A. nidulans genes, and 13 GSs were of DNA or genes encoding cytoplasmic or organellar proteins. This pattern is similar to those in the human HepG2 cell line and in human colonic mucosa, although very few genes for nuclear proteins and for protein synthesis were in A. nidulans.

• Mycobiology
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• v.43 no.1
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• pp.31-36
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• 2015

We have previously isolated ${\varepsilon}$-COP, the ${\alpha}$-COP interactor in COPI of Aspergillus nidulans, by yeast two-hybrid screening. To understand the function of ${\varepsilon}$-COP, the $aneA^+$ gene for ${\varepsilon}$-COP/AneA was deleted by homologous recombination using a gene-specific disruption cassette. Deletion of the ${\varepsilon}$-COP gene showed no detectable changes in vegetative growth or asexual development, but resulted in decrease in the production of the fruiting body, cleistothecium, under conditions favorable for sexual development. Unlike in the budding yeast Saccharomyces cerevisiae, in A. nidulans, over-expression of ${\varepsilon}$-COP did not rescue the thermo-sensitive growth defect of the ${\alpha}$-COP mutant at $42^{\circ}C$. Together, these data show that ${\varepsilon}$-COP is not essential for viability, but it plays a role in fruiting body formation in A. nidulans.

• Korean Journal of Microbiology
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• v.38 no.2
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• pp.103-106
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• 2002

Direct PCR from intact fungal cells is not readily suitable to all fungi mainly because of difficulties in rupturing the cell walls. Microwave irradiation has been proven to be useful in fungal DNA extraction protocol. Here we describe a fast template preparation method for PCR amplification from Aspefillus nidulans conidiospores using microwave irradiation. We optimized the duration far microwave irradiation, and the amount of template DNA for PCR. Amplification from samples prepared in this manner was so efficient that we could get PCR products with size enough to identify transformants. We believe that this is a time-saving procedure for screening true transformants of A. nidulans.